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Liver-expressed antimicrobial peptide 2 (LEAP2) is a small peptide, which is consisted of signal peptide, pro-peptide and the bioactive mature peptide. Mature LEAP2 is an antibacterial peptide with four highly conserved cysteines forming two intramolecular disulfide bonds. Chionodraco hamatus, an Antarctic notothenioid fish that lives in the coldest water, has white blood unlike most fish of the world. In this study, the LEAP2 coding sequence was cloned from C. hamatus, including a 29 amino acids signal peptide and mature peptide of 46 amino acids. High levels of LEAP2 mRNA were detected in the skin and liver. Mature peptide was obtained by chemical synthesis in vitro, displayed selective antimicrobial activities against Escherichia coli, Aeromonas hydrophila, Staphylococcus aureus and Streptococcus agalactiae. Liver-expressed antimicrobial peptide 2 showed bactericidal activity by destroying the cell membrane integrity and robustly combined with bacterial genomic DNA. In addition, overexpression of the Tol-LEAP2-EGFP in zebrafish larva showed stronger antimicrobial activity in C. hamatus than in zebrafish, accompanied by lower bacterial load and expression of pro-inflammatory factors. This is the first demonstration of the antimicrobial activity of LEAP2 from C. hamatus, which is of useful value in improving resistance to pathogens., (© 2023 John Wiley & Sons Ltd.)

The Antarctic icefish, a family (Channichthyidae) of teleosts within the perciform suborder Notothenioidei, are the only known vertebrates without oxygen-transporting haemoglobins and that are largely devoid of circulating erythrocytes. To elucidate the evo-devo mechanisms underpinning the suppressed erythropoiesis in the icefish, we conducted comparative studies on the transcriptomes and microRNAomes of the primary haematopoietic tissues between an icefish (Chionodraco hamatus) and two red-blooded notothenioids (Trematomus bernacchii and Gymnodraco acuticeps). We identified substantial remodelling of the haematopoietic programs in the icefish through which erythropoiesis is selectively suppressed. Experimental verification showed that erythropoietic suppression in the icefish may be attributable to the upregulation of TGF-β signalling, which coincides with reductions in multiple transcription factors essential for erythropoiesis and the upregulation of hundreds of microRNAs, the majority (> 80%) of which potentially target erythropoiesis regulating factors. Of the six microRNAs selected for verification, three miRNAs (miR-152, miR-1388 and miR-16b) demonstrated suppressive functions on GATA1 and ALAS2, which are two factors important for erythroid differentiation, resulting in reduced numbers of erythroids in microinjected zebra fish embryos. Codon substitution analyses of the genes of the TGF-β superfamily revealed signs of positive selection in TGF-β1 and endoglin in the lineages leading to Antarctic notothenioids. Both genes are previously known to function in erythropoietic suppression. These findings implied a general trend of erythropoietic suppression in the cold-adapted notothenioid lineages through evolutionary modulation of the multi-functional TGF-β signalling pathway. This trend is more pronounced in the haemoglobin-less icefish, which may pre-emptively hinder the otherwise defective erythroids from production., (© 2015 John Wiley & Sons Ltd.)